Cobas modular system

Blood glucose and insulin were analysed with a Cobas Modular system (Roche Diagnostics, Risch, Switzerland) at the Clinical Chemistry Laboratory, Sahlgrenska University Hospital (accredited in accordance with the International Standard ISO 15189:2007). CSF insulin was analyzed with a double-antibody radioimmunoassay (Linco Research) at the Department of Clinical Science, Lund University. Leptin and adiponectin were measured with ELISA kits (R&D Systems) in the Clinical Neurochemistry Laboratory at Sahlgrenska University Hospital Mölndal. ELISA plates were read on a Vmax plate reader, and concentrations were determined with Softmax software (Molecular Devices). Insulin was measured in undiluted samples and adiponectin in 100-fold diluted samples. For leptin analysis, CSF samples were diluted 2-fold and serum samples 100-fold. HOMA-IR was calculated as (fasting glucose × fasting insulin)/22.5 and HOMA-B as (20 × fasting insulin)/(fasting glucose − 3.5) [31 (link)]. GDF15 concentration was measured with Human GDF-15 Quantikine Elisa Kit (R&D Systems, Minneapolis, MN, USA). Serum GDF15 samples during and after pregnancy were diluted 1:64 and 1:4, respectively. CSF GDF15 samples were diluted 1:2. The intra- and inter-assay coefficients of variation (CVs) for GDF15 measurements were 1.7% and 7%, respectively.

Serum creatine kinase (CK) concentrations were analyzed by a spectrophotometric method according to a previously described method (31 (link)). IL-6 and TNF-α were assayed using ELISA kits from R&D Systems, Minneapolis, MN, USA (catalog No. HS600B and HSTA00C, respectably). Variation coefficient (CV) for CK, IL-6 and TNF-α is 3–4, 9, and 16%, respectively (31 (link), 32 (link)). Glucose, insulin, cholesterol (total, low-density lipoprotein [LDL], high-density lipoprotein [HDL]), triglycerides (TG) and C-reactive protein (CRP) were routinely analyzed at the accredited Clinical Chemistry Laboratory, Sahlgrenska University Hospital (International Standard ISO 15189:2007) using a Cobas Modular system (Roche Diagnostics, Risch, Switzerland). CV for insulin was 10%, TG 4%, cholesterol 3%, CRP 3–4% and glucose 3%. Homeostatic model of insulin resistance (HOMA-IR) was calculated as (fasting glucose × fasting insulin)/22.5 (33 (link)). Two fasting blood samples were excluded from analysis due to participants not complying with the fasting protocol.